SPP-ARV-825-mediated cells apoptosis in vitro. (A) Flow cytometry detection of GL261 cells apoptosis after different treatments for 48 h. (B) Statistical analysis of apoptosis cells (n = 3). (C) Flow cytometry analysis of mitochondrial membrane potential levels of GL261 cells stained with JC-1 after treatment with 400 ng/mL SPP-ARV-825 for 48 h. Untreated cells were used as the control. (D) Statistical analysis of the red/green fluorescence signal ratio of JC-1 (n = 3). (E) Expressions of apoptosis related proteins (Bax, MCL-1, BCL-2, pro-caspase 8, cleaved caspase 8, pro-caspase 3, cleaved caspase 3, PARP) and the loading control GAPDH were determined by Western blot analysis after treatment of GL261 cells with different concentrations of SPP-ARV-825. Data are presented as mean ± SEM. ∗∗∗∗P < 0.0001.